A KDELR-mediated ER-retrieval system modulates mitochondrial functions via the unfolded protein response in fission yeast.

KDELR (Erd2 [ER retention defective 2] in yeasts) is a receptor protein that retrieves endoplasmic reticulum (ER)-resident proteins from the Golgi apparatus. However, the role of the KDELR-mediated ER-retrieval system in regulating cellular homeostasis remains elusive. Here, we show that the absence of Erd2 triggers the unfolded protein response (UPR) and enhances mitochondrial respiration and reactive oxygen species in an UPR-dependent manner in the fission yeast Schizosaccharomyces pombe. Moreover, we perform transcriptomic analysis and find that the expression of genes related to mitochondrial respiration and the tricarboxylic acid cycle is upregulated in a UPR-dependent manner in cells lacking Erd2. The increased mitochondrial respiration and reactive oxygen species production is required for cell survival in the absence of Erd2. Therefore, our findings reveal a novel role of the KDELR-Erd2-mediated ER-retrieval system in modulating mitochondrial functions and highlight its importance for cellular homeostasis in the fission yeast.

Whole-genome sequencing and transcriptome-characterized mechanism of streptomycin resistance in Vibrio parahaemolyticus O10: K4.

Streptomycin resistance in V. parahaemolyticus has been widespread in both clinical and environmental isolates. Therefore, it is of great significance to characterize the mechanism of streptomycin resistance in V. parahaemolyticus. O10:K4 has emerged and becoming the new dominant serotype since 2020. In this study, we isolated a total of 36 strains of V. parahaemolyticus O10:K4 from 2020 to 2022 and found that more than half of them were resistant to streptomycin. We obtained streptomycin resistant and sensitive strains by detecting the resistance profiles. Whole-genome sequencing showed that VP_RS10735 and VP_RS05605 were the predominant mutations in streptomycin resistant O10:K4 clinical isolates. In addition, this study provided global insight into the characteristics of the transcriptome signature of streptomycin resistance, revealing that efflux transporters play a key role in streptomycin resistance. Finally, we found that streptomycin resistant strain was more virulent than sensitive strain. The results of this study should advance our understanding of the mechanisms of aminoglycoside resistance.

SARS-CoV-2 genomic characterization and evolution in China.

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) affected global health worldwide due to its high contagiousness. During the viral spread, many mutations occurred within the virus genome. China has adopted nonpharmaceutical intervention (NPI) to contain COVID-19 outbreaks. In order to understand the evolution and genomic variation of SARS-CoV-2 in China under this policy, a total of 524 sequences downloaded from Global Initiative on Sharing All Influenza Data (GISAID) between 2019 and 2022 were included in this study. The time-scaled evolutionary analysis showed that these sequences clustered in three groups (Group A-C). Group B and C accounted for the majority of the sequences whose divergence times were around 2020 and distributed in multiple regions. Group A was mainly composed of G variants, which were mainly isolated from several regions. Moreover, we found that 191 sites had mutations with no less than 3 times, including 30 amino acids that were deleted. Finally, we found that spike and nucleocapsid genes underwent positive selection evolution, indicating that the mutations within spike and nucleocapsid genes increased the SARS-CoV-2 contagiousness. Hence, this study preliminarily elucidates the evolutionary characteristics and genomic mutations of SARS-CoV-2 under the implementation of the NPI policy in China, providing scientific basis for further understanding the control effect of the NPI policy on the epidemic.

Genomic characterization of Campylobacter isolates in Huzhou, China.

Campylobacter is a major foodborne pathogen that causes outbreaks and sporadic gastrointestinal disease, creating a serious disease burden. Campylobacter strains isolated from diarrhea cases (n = 11) and raw poultry meat products (n = 2) in Huzhou, including 11 Campylobacter jejuni and 2 Campylobacter coli strains, were subjected to virulence gene, drug resistance gene, genetic correlation, antibiotic resistance, and multilocus sequence typing (MLST) analyses. The 13 Campylobacter isolates were divided into 12 sequence types (STs), one of which was a new ST. The isolated strains contain multiple virulence-related genes. Drug sensitivity analysis showed that the resistance rate of the 13 isolates to nalidixic acid, ciprofloxacin, and tetracycline was 92.3%. Genome sequencing indicated that all 11 strains of C. jejuni carried the tet(O) and blaOXA resistance genes, and 2 strains of C. coli carried multiple drug resistance genes. Phylogenetic analysis based on core-genome single-nucleotide polymorphisms indicated that the 11 C. jejuni isolates from diarrhea patients and food sources are not closely phylogenetically related.

Mitochondrial fusion and fission are required for proper mitochondrial function and cell proliferation in fission yeast.

Mitochondria form a branched tubular network in many types of cells, depending on a balance between mitochondrial fusion and fission. How mitochondrial fusion and fission are involved in regulating mitochondrial function and cell proliferation is not well understood. Here, we dissected the roles of mitochondrial fusion and fission in mitochondrial function and cell proliferation in fission yeast. We examined mitochondrial membrane potential by staining cells with DiOC6 and assessed mitochondrial respiration by directly measuring oxygen consumption of cells with a dissolved oxygen respirometer. We found that defects in mitochondrial fission or fusion reduce mitochondrial membrane potential and compromise mitochondrial respiration while the absence of both mitochondrial fusion and fission restores wild type-like respiration, normal membrane potential, and tubular networks of mitochondria. Moreover, we found that the absence of either mitochondrial fission or fusion prolongs the cell cycle and that the absence of both mitochondrial fusion and fission significantly delays cell cycle progression after nitrogen replenishment. The prolonged/delayed cell cycle is likely due to the deregulation of Cdc2 activation. Hence, our work not only establishes an intimate link between mitochondrial morphology and function but also underscores the importance of mitochondrial dynamics in regulating the cell cycle.

Emr1 regulates the number of foci of the endoplasmic reticulum-mitochondria encounter structure complex.

The endoplasmic reticulum-mitochondria encounter structure (ERMES) complex creates contact sites between the endoplasmic reticulum and mitochondria, playing crucial roles in interorganelle communication, mitochondrial fission, mtDNA inheritance, lipid transfer, and autophagy. The mechanism regulating the number of ERMES foci within the cell remains unclear. Here, we demonstrate that the mitochondrial membrane protein Emr1 contributes to regulating the number of ERMES foci. We show that the absence of Emr1 significantly decreases the number of ERMES foci. Moreover, we find that Emr1 interacts with the ERMES core component Mdm12 and colocalizes with Mdm12 on mitochondria. Similar to ERMES mutant cells, cells lacking Emr1 display defective mitochondrial morphology and impaired mitochondrial segregation, which can be rescued by an artificial tether capable of linking the endoplasmic reticulum and mitochondria. We further demonstrate that the cytoplasmic region of Emr1 is required for regulating the number of ERMES foci. This work thus reveals a crucial regulatory protein necessary for ERMES functions and provides mechanistic insights into understanding the dynamic regulation of endoplasmic reticulum-mitochondria communication.

Centromere targeting of Mis18 requires the interaction with DNA and H2A-H2B in fission yeast.

Faithful chromosome segregation during mitosis requires the correct assembly of kinetochore on the centromere. CENP-A is a variant of histone H3, which specializes the centromere region on chromatin and mediates the kinetochore assembly. The Mis18 complex plays a critical role in initiating the centromere loading of the newly-synthesized CENP-A. However, it remains unclear how Mis18 complex (spMis18, spMis16 and spMis19) is located to the centromere to license the recruitment of Cnp1CENP-A in Schizosaccharomyces pombe. We found that spMis18 directly binds to nucleosomal DNA through its extreme C-terminus and interacts with H2A-H2B dimer via the acidic region on the surface of its Yippee-like domain. Live-cell imaging confirmed that mutation of the acidic region and deletion of the extreme C-terminus significantly impairs the localization of spMis18 and Cnp1 to the centromere and delays chromosome segregation during mitosis. Our findings illustrate that the interaction of spMis18 with histone H2A-H2B and DNA plays important roles in the recruitment of spMis18 and Cnp1 to the centromere in fission yeast.

Klp2 and Ase1 synergize to maintain meiotic spindle stability during metaphase I.

The spindle apparatus segregates bi-oriented sister chromatids during mitosis but mono-oriented homologous chromosomes during meiosis I. It has remained unclear if similar molecular mechanisms operate to regulate spindle dynamics during mitosis and meiosis I. Here, we employed live-cell microscopy to compare the spindle dynamics of mitosis and meiosis I in fission yeast cells and demonstrated that the conserved kinesin-14 motor Klp2 plays a specific role in maintaining metaphase spindle length during meiosis I but not during mitosis. Moreover, the maintenance of metaphase spindle stability during meiosis I requires the synergism between Klp2 and the conserved microtubule cross-linker Ase1, as the absence of both proteins causes exacerbated defects in metaphase spindle stability. The synergism is not necessary for regulating mitotic spindle dynamics. Hence, our work reveals a new molecular mechanism underlying meiotic spindle dynamics and provides insights into understanding differential regulation of meiotic and mitotic events.

Glucose starvation induces mitochondrial fragmentation depending on the dynamin GTPase Dnm1/Drp1 in fission yeast.

Mitochondria undergo morphological and dynamic changes in response to environmental stresses. Few studies have focused on addressing mitochondrial remodeling under stress. Using the fission yeast Schizosaccharomyces pombe as a model organism, here we investigated mitochondrial remodeling under glucose starvation. We employed live-cell microscopy to monitor mitochondrial morphology and dynamics of cells in profusion chambers under glucose starvation. Our results revealed that mitochondria fragment within minutes after glucose starvation and that the dynamin GTPase Dnm1 is required for promoting mitochondrial fragmentation. Moreover, we found that glucose starvation enhances Dnm1 localization to mitochondria and increases the frequency of mitochondrial fission but decreases PKA activity. We further demonstrate that low PKA activity enhances glucose starvation-induced mitochondrial fragmentation, whereas high PKA activity confers resistance to glucose starvation-induced mitochondrial fragmentation. Moreover, we observed that AMP-activated protein kinase is not involved in regulating mitochondrial fragmentation under glucose starvation. Of note, glucose starvation-induced mitochondrial fragmentation was associated with enhanced reactive oxygen species production. Our work provides detailed mechanistic insights into mitochondrial remodeling in response to glucose starvation.

Automated morphometry toolbox for analysis of microscopic model organisms using simple bright-field imaging.

Model organisms with compact genomes, such as yeast and C aenorhabditis elegans, are particularly useful for understanding organism growth and life/cell cycle. Organism morphology is a critical parameter to measure in monitoring growth and stage in the life cycle. However, manual measurements are both time consuming and potentially inaccurate, due to variations among users and user fatigue. In this paper we present an automated method to segment bright-field images of fission yeast, budding yeast, and C. elegans roundworm, reporting a wide range of morphometric parameters, such as length, width, eccentricity, and others. Comparisons between automated and manual methods on fission yeast reveal good correlation in size values, with the 95% confidence interval lying between -0.8 and +0.6 µm in cell length, similar to the 95% confidence interval between two manual users. In a head-to-head comparison with other published algorithms on multiple datasets, our method achieves more accurate and robust results with substantially less computation time. We demonstrate the method's versatility on several model organisms, and demonstrate its utility through automated analysis of changes in fission yeast growth due to single kinase deletions. The algorithm has additionally been implemented as a stand-alone executable program to aid dissemination to other researchers.

Septins regulate the equatorial dynamics of the separation initiation network kinase Sid2p and glucan synthases to ensure proper cytokinesis.

Septins generally function as scaffolds and as cortical barriers to restrict the diffusion of membrane proteins. In the fission yeast Schizosaccharomyces pombe, septins form a ring structure at the septum after spindle breakdown during the constriction of the contractile actomyosin ring (CAR) and serve as a scaffold to recruit glucanases to mediate ultimate daughter cell separation. Despite this, it remains unclear if septins play any significant roles before the cell separation during cytokinesis. Employing live cell microscopy, we carefully examined SIN (Septation Initiation Network) signaling and glucan synthases, two key factors ensuring proper function of the CAR. In the absence of the core septin component Spn1p, the formation of a compact CAR is advanced and the CAR constriction rate is slightly but significantly decreased. Moreover, the SIN kinase Sid2p and the glucan synthases Bgs1p and Ags1p form an equatorial ring quite prematurely, but their maintenance at the equatorial region is diminished spn1Δ cells. These findings suggest that septins act as key players in an accurate establishment and the maintenance of CAR by orchestrating the equatorial dynamics of Sid2p and glucan synthases. Hence, this work demonstrates that, in addition to their function during ultimate cell septation, septins have important roles in regulating earlier cytokinetic events, including CAR assembly and constriction, SIN signaling, and the cortical dynamics of the glucan synthases.